Biodegradable metal alloys

ABSTRACT

The invention relates to biodegradable, metal alloy-containing compositions, methods for their preparation and applications for their use. The compositions include magnesium and other components, such as yttrium, calcium, silver, cerium, and zirconium; or zinc, silver, cerium, and zirconium; or aluminum, zinc, calcium, manganese, silver, yttrium; or strontium, calcium, zinc. The compositions are prepared by vacuum induction/crucible melting together the components and casting the melted mixture in a preheated mild steel/copper mold. In certain embodiments, the compositions of the invention are particularly useful for forming medical devices for implantation into a body of a patient.

The invention was made with government support under EEC-0812348 awardedby the National Science Foundation. The government has certain rights inthe invention.

FIELD OF THE INVENTION

The invention relates to metal alloy-containing compositions andarticles, and methods for their preparation. The invention isparticularly suitable for use in fabricating biodegradable materials andmedical devices for implantation into a body of a patient, such as forexample, orthopedic, craniofacial and cardiovascular implant devices.

BACKGROUND OF THE INVENTION

Metallic implant devices, such as plates, screws, nails and pins arecommonly used in the practice of orthopedic, craniofacial andcardiovascular implant surgery. Furthermore, metallic stents are alsoimplanted into a body of a patient to support lumens, for example,coronary arteries. Most of these metallic implant devices which arecurrently used are constructed of stainless steel, cobalt-chromium(Co—Cr) or titanium alloys. Advantageously, these materials ofconstruction exhibit good biomechanical properties. However,disadvantageously, implant devices constructed of these materials do notdegrade over a period of time. Thus, surgery may be required when thereis no longer a medical need for the implant device and when, for variousreasons, it may be desired to remove the implant device from a body of apatient. For example, in certain instances, such as pediatricapplications, there may be a concern that if an implant device is notremoved, it may eventually be rejected by the body and causecomplications for the patient. Thus, it would be advantageous for: (i)the implant device to be constructed of a material that is capable ofdegrading over a period of time, (ii) for the implant device to dissolvein a physiological environment such that it would not remain in the bodywhen there is no longer a medical need for it, and (iii) surgery not tobe required to remove the implant device from the body of the patient.

Currently, biomaterials used for orthopedic, craniofacial andcardiovascular applications are primarily chosen based on their abilityto withstand cyclic load-bearing. Metallic biomaterials in particularhave appropriate properties such as high strength, ductility, fracturetoughness, hardness, corrosion resistance, formability, andbiocompatibility to make them attractive for most load bearingapplications. The most prevalent metals for load-bearing applicationsare stainless steels, Ti, and Co—Cr based alloys, though theirstiffness, rigidity, and strength far exceed those of natural bone.Their elastic modulus differs significantly from bone, causingstress-shielding effects that may lead to reduced loading of bone—withthis decrease in stimulation resulting in insufficient new bone growthand remodeling, decreasing implant stability. Current metallicbiomaterials also suffer from the risk of releasing toxic metallic ionsand particles through corrosion or wear causing implant site immuneresponse. They may also lead to hypersensitivity, growth restriction(most significantly for pediatric implants), implant migration, andimaging interference. Due to these complications, it is estimated that10% of patients will require a second operation for the removal ofpermanent metallic plates and screws, exposing patients to additionalrisks, and increasing surgical time and resources.

Based on at least these issues, there is a desire to design and developa new class of load-bearing biomaterials with the goal of providingadequate support while the bone is healing that harmlessly degrades overtime.

To avoid complications associated with permanent fixation implants,degradable biomaterials have recently been developed. However,resorbable polymer fixation plates and screws are relatively weaker andless rigid compared to metals, and have demonstrated local inflammatoryreactions. For example, biodegradable materials which are currently usedin the construction of implant devices include polymers, such aspolyhydroxy acids, polylactic acid (PLA), polyglycolic acid (PGA), andthe like. These materials, however, have been found to exhibitrelatively poor strength and ductility, and have a tendency to reactwith human tissue which can limit bone growth.

Magnesium alloys have recently emerged as a new class of biodegradablematerials for orthopedic applications with more comparable properties tonatural bone. Magnesium is known to be a non-toxic metal element thatdegrades in a physiological environment and therefore, may be considereda suitable element for use in constructing biodegradable implantdevices. Magnesium is attractive as a biomaterial for several reasons.It is very lightweight, with a density similar to conical bone, and muchless than stainless steel, titanium alloys, and Co—Cr alloys. Theelastic modulus of magnesium is much closer to natural bone compared toother commonly used metallic implants, thus reducing the risk of stressshielding. Magnesium is also essential to human metabolism, is acofactor for many enzymes, and stabilizes the structures of DNA and RNA.Most importantly, magnesium degrades to produce a soluble, non-toxiccorrosion hydroxide product which is harmlessly excreted through urine.Unfortunately, accelerated corrosion of magnesium alloys may lead toaccumulation of hydrogen gas pockets around the implant as well asinsufficient mechanical performance and implant stability throughout thedegradation and tissue healing process. The degradation of magnesium ina physiological environment yields magnesium hydroxide and hydrogen gas.This process is known in the art as magnesium corrosion. The hydrogengas produced in the body of the patient as a result of magnesiumcorrosion can produce complications because the ability of the humanbody to absorb or release hydrogen gas is limited.

The various biodegradable metallic alloys known in the art may exhibitlow biocompatibility and/or high corrosion rates, which render thesematerials unsuitable for use in medical applications, such as implantdevices. Further, compositions of matter for use as implant devicesshould not include toxic elements, such as zinc and aluminum, or atleast include these elements only in non-toxic amounts. Moreover, thecomposition should exhibit a corrosion rate that is suitable forimplantation in a physiological environment, i.e., a body of a patient.

In the field of biomedical applications, there is a desire to developbiodegradable metal alloy-containing implant materials having goodcompressive strength with improved corrosion resistance andbiocompatibility. Further, it is desirable to control the corrosionresistance and the hydrogen evolution therefrom, which is associatedwith the presence of magnesium in a physiological environment.

SUMMARY OF THE INVENTION

In one aspect, the invention provides a biodegradable, metalalloy-containing composition including from about 0.5 weight percent toabout 4.0 weight percent of yttrium, from greater than zero to about 1.0weight percent of calcium, from about 0.25 weight percent to about 1.0weight percent of zirconium, and a balance of magnesium, based on thetotal weight of the composition. In certain embodiments, the metalalloy-containing composition can include about 1.0 weight percent ofyttrium. In another embodiment, the metal alloy-containing compositioncan include about 1.0 weight percent of calcium. In still anotherembodiment, the metal alloy-containing composition can include less thanabout 0.5 weight percent of zirconium.

In another aspect, the invention provides a biodegradable, metalalloy-containing composition including from about 1.0 weight percent toabout 6.0 weight percent of zinc, from greater than zero to about 1.0weight percent of zirconium, and a balance of magnesium, based on thetotal weight of the composition. In certain embodiments, the metalalloy-containing composition can include about 4.0 weight percent ofzinc. In another embodiment, the metal alloy-containing composition caninclude less than about 0.5 weight percent of zirconium.

In another aspect, the invention provides a method of preparing abiodegradable, metal alloy-containing composition including melting fromabout 0.5 weight percent to about 4.0 weight percent of yttrium, fromgreater than zero to about 1.0 weight percent of calcium, from about0.25 weight percent to about 1.0 weight percent of zirconium, and abalance of magnesium, based on the total weight of the composition, toobtain a melted mixture and casting the melt mixture to obtain saidbiodegradable, metal alloy-containing composition. In certainembodiments, the method can include melting about 1.0 weight percent ofyttrium. In another embodiment, the method can include melting about 1.0weight percent of calcium. In still another embodiment, the method caninclude melting less than about 0.5 weight percent of zirconium.

In another aspect, the invention provides a method of preparing abiodegradable, metal alloy-containing composition including melting fromabout 1.0 weight percent to about 6.0 weight percent of zinc, fromgreater than zero to about 1.0 weight percent of zirconium, and abalance of magnesium, based on the total weight of the composition, toobtain a melted mixture and casting the melt mixture to obtain saidbiodegradable, metal alloy-containing composition. In certainembodiments, the method can include melting about 4.0 weight percent ofzinc. In another embodiment, the method can include melting less thanabout 0.5 weight percent of zirconium.

In yet another aspect, the invention includes a biodegradable, metalalloy-containing article including a magnesium-containing compositionincluding from about 0.5 weight percent to about 4.0 weight percent ofyttrium, from greater than zero to about 1.0 weight percent of calcium,from about 0.25 weight percent to about 1.0 weight percent of zirconium,and a balance of magnesium, based on the total weight of thecomposition. In certain embodiments, the magnesium-containingcomposition can include about 1.0 weight percent of yttrium. In anotherembodiment, the magnesium-containing composition can include about 1.0weight percent of calcium. In still another embodiment, themagnesium-containing composition can include less than about 0.5 weightpercent of zirconium.

In yet another aspect, the invention includes a biodegradable, metalalloy-containing article including a magnesium-containing compositionincluding from about 1.0 weight percent to about 6.0 weight percent ofzinc, from greater than zero to about 1.0 weight percent of zirconium,and a balance of magnesium, based on the total weight of thecomposition. In certain embodiments, the magnesium-containingcomposition can include about 4.0 weight percent of zinc. In anotherembodiment, the magnesium-containing composition can include less thanabout 0.5 weight percent of zirconium.

In still another aspect, the invention includes a biodegradable, metalalloy-containing medical device including a magnesium-containingcomposition including about 0.5 weight percent to about 4.0 weightpercent of yttrium, from greater than zero to about 1.0 weight percentof calcium, from about 0.25 weight percent to about 1.0 weight percentof zirconium, and a balance of magnesium, based on the total weight ofthe composition. In certain embodiments, the magnesium-containingcomposition can include about 1.0 weight percent of yttrium. In anotherembodiment, the magnesium-containing composition can include about 1.0weight percent of calcium. In still another embodiment, themagnesium-containing composition can include less than about 0.5 weightpercent of zirconium. In certain embodiments, this medical device can beimplantable in a body of a patient. In another embodiment, the medicaldevice can be an orthopedic device. In yet another embodiment, themedical device can be a craniofacial device. In still anotherembodiment, the medical device can be a cardiovascular device.

In still another aspect, the invention includes a biodegradable, metalalloy-containing medical device including a magnesium-containingcomposition including about 1.0 weight percent to about 6.0 weightpercent of zinc, from greater than zero to about 1.0 weight percent ofzirconium, and a balance of magnesium, based on the total weight of thecomposition. In certain embodiments, the magnesium-containingcomposition can include about 4.0 weight percent of zinc. In anotherembodiment, the magnesium-containing composition can include less thanabout 0.5 weight percent of zirconium. In certain embodiments, thismedical device can be implantable in a body of a patient. In anotherembodiment, the medical device can be an orthopedic device. In yetanother embodiment, the medical device can be a craniofacial device. Instill another embodiment, the medical device can be a cardiovasculardevice.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The invention relates to novel, biodegradable metal alloy-containingcompositions. Further, the invention relates to articles, such asmedical devices for implantation into a body of a patient, which areconstructed or fabricated from the biodegradable metal alloy-containingcompositions of the invention. Moreover, the invention relates tomethods of preparing these biodegradable, metal alloy-containingcompositions and articles for use in medical applications, such as butnot limited to, orthopedic, craniofacial and cardiovascular surgery.

In addition to the biodegradability of the metal alloy-containingcompositions of the invention, these compositions include at least oneof the following characteristics: biocompatibility, corrosionresistance, cell attachment, viability and mechanical strength, whichmake them suitable for use as implant devices in a body of a patient.

In certain embodiments, the biodegradable, metal alloy-containingcompositions of the invention are based on the presence of magnesium.The amount of magnesium and additional components are selected such thatthe compositions exhibit the characteristics identified herein. Forexample, components and their amounts are selected such that thecompositions exhibit corrosion resistance in the presence of water andsimulated body fluids which allow the compositions to be suitable for invitro use, for example, in a physiological environment, such as a bodyof a patient.

In other embodiments, the biodegradable, metal alloy-containingcompositions of the invention are prepared using selected components inspecified amounts such that the compositions exhibit corrosionresistance with minimal or no evolution of hydrogen gas. The evolutionof hydrogen, such as, hydrogen bubbles may result in complicationswithin a body of a patient.

This invention includes controlling the corrosion rate and improvingmechanical properties of magnesium alloys through the introduction ofalloying elements and processing conditions. Magnesium corrosion andmechanical properties are strongly affected by alloying elements in thesolid solution.

In certain embodiments, the biodegradable, metal alloy-containingcompositions of the invention include the following components: yttrium,calcium, zirconium and magnesium. The amount of each of these componentsin the compositions can vary. In general, the amounts of each of thesecomponents are selected in order that the resulting compositions arewithin acceptable non-toxic limits such that the compositions aresufficiently biocompatible for implantation into a body of a patient,and are degradable over a period of time so that the implantation devicedoes not remain in the body of the patient for prolonged periods oftime, e.g., not beyond the period of time when there is a medical needfor the implantation device. An implantation device fabricated inaccordance with the invention will degrade and preferably completelydissolve within an acceptable time frame. For example, an implant devicefabricated in accordance with the invention can serve as filler orsupport material during a bone healing process and following completionof this process, the implant device will degrade within an acceptabletime period and therefore, will not remain in the body for a prolongedperiod of time. The acceptable non-toxic limits and the acceptable timeframe for degradation can vary and can depend on particular physical andphysiological characteristics of the patient, the particular in vivesite of the implantation device, and the particular medical use of theimplantation device.

In certain embodiments, the composition of the invention includes fromabout 0.5 weight percent to about 4.0 weight percent of yttrium, fromgreater than zero to about 1.0 weight percent of calcium, from about0.25 weight percent to about 1.0 weight percent of zirconium, and theremainder or balance being magnesium based on the total weight of thecomposition. In other embodiments, the composition can include about 1.0weight percent of yttrium or about 4.0 weight percent of yttrium. In yetother embodiments, the composition can include about 1.0 weight percentof calcium or about 0.6 weight percent of calcium. In still otherembodiments, the composition can include less than about 0.5 weightpercent of zirconium, or about 0.4 weight percent of zirconium.

Without intending to be bound by any particular theory, it is believedthat the presence of yttrium contributes to the improved mechanicalstrength and corrosion resistance of the biodegradable, metalalloy-containing compositions. Calcium is used in a low quantity toprevent oxidation during the casting of the alloy. Zirconium is known toact as a grain refiner and is used to improve mechanical properties ofthe compositions.

In another embodiment of the invention, the biodegradable, metalalloy-containing compositions of the invention include the followingcomponents: zinc, zirconium and magnesium. The amount of each of thesecomponents in the compositions can vary. As previously indicated, ingeneral, the amounts of each of these components are selected in orderthat the resulting compositions are within acceptable non-toxic limitsand are degradable over an acceptable period of time. In certainembodiments, the composition of the invention includes from about 1.0weight percent to about 6.0 weight percent of zinc, from greater thanzero to about 1.0 weight percent of zirconium, and the remainder orbalance being magnesium based on the total weight of the composition. Inanother embodiment, the composition can include about 4.0 weight percentof zinc. In still another embodiment, the composition can include lessthan about 0.5 weight percent of zirconium.

As described previously herein, the use of magnesium-containingcompositions in a physiological environment results in the evolution orproduction of hydrogen gas. The degradation of magnesium involves aprocess (i.e., a corrosion process) in which hydrogen is released. Inthe invention, the amount of magnesium is specified such that thecorrosion rate corresponds to a rate of hydrogen formation which isacceptable such that large amounts of hydrogen bubbles do not form andaccumulate within a body of a patient.

In certain embodiments, the amounts of yttrium, calcium, zirconium andmagnesium are specified and adjusted such as to control at least one ofcorrosion resistance, biodegradation, biocompatibility, toxicity, cellattachment, mechanical strength and flexibility. In other embodiments,the amounts of zinc, zirconium and magnesium are specified and adjustedsuch as to control at least one of corrosion resistance, biodegradation,biocompatibility, toxicity, cell attachment, mechanical strength andflexibility.

Further, in certain embodiments, other compounds may be added to impartadditional characteristics and properties to the resultingbiodegradable, metal alloy-containing compositions. For example, silvermay be added to provide anti-microbial properties.

Non-limiting examples of medical devices in which the compositions andarticles of the invention can be used include, but are not limited toplates, meshes, staples, screws, pins, tacks, rods, suture anchors,tubular mesh, coils, x-ray markers, catheters, endoprostheses, pipes,shields, bolts, clips or plugs, dental implants or devices, graftdevices, bone-fracture healing devices, bone replacement devices, jointreplacement devices, tissue regeneration devices, cardiovascular stents,intercranial aneurism device, tracheal stents, nerve guides, surgicalimplants and wires. In a preferred embodiment, the medical devicesinclude fixation bone plates and screws, temporamandibular joints,cardiovascular stents, and nerve guides.

The medical devices described herein can have at least one activesubstance attached thereto. The active substance can be either attachedto the surface or encapsulated within. As used herein, the term “activesubstance” describes a molecule, compound, complex, adduct and/orcomposite that exhibits one or more beneficial activities such astherapeutic activity, diagnostic activity, biocompatibility, corrosion,and the like. Active substances that exhibit a therapeutic activity caninclude bioactive agents, pharmaceutically active agents, drugs and thelike. Non-limiting examples of bioactive agents that can be incorporatedin the compositions, articles and devices of the invention include, butare not limited to, bone growth promoting agents such as growth factors,drugs, proteins, antibiotics, antibodies, ligands. DNA, RNA, peptides,enzymes, vitamins, cells and the like, and combinations thereof.

It is contemplated that additional components may be added to thebiodegradable, metal alloy-containing compositions of the inventionprovided that the non-toxicity and biodegradability of the compositionsis maintained within acceptable limits. The additional components can beselected from a wide variety known in the art and can include one ormore of cerium, aluminum, strontium, manganese and silver.

In certain embodiments, aluminum is present in an amount of from about1.0 to 9.0 weight percent based on total weight of the composition. Inother embodiments, the aluminum is present in an amount of about 2.0weight percent based on total weight of the composition.

In certain embodiments, manganese is present in an amount of from about0.1 to about 1.0 weight percent based on total weight of thecomposition. In other embodiments, the manganese is present in an amountof about 0.2 weight percent based on total weight of the composition.

In certain embodiments, silver is present in an amount of from about0.25 to about 1.0 weight percent based on total weight of thecomposition. In other embodiments, the silver is present in an amount ofabout 0.25 weight percent based on total weight of the composition.

In certain embodiments, cerium is present in an amount of from about 0.1to about 1.0 weight percent based on total weight of the composition. Inother embodiments, the cerium is present in an amount of about 0.5weight percent based on total weight of the composition.

In certain embodiments, strontium is present in an amount of from about1.0 to about 4.0 weight percent based on total weight of thecomposition. In other embodiments, the strontium can be present in anabout of 3.0 weight percent.

In one embodiment, the biodegradable, metal-alloy containing compositionincludes from about 0.5 weight percent to about 4.0 weight percent ofytrrium, from greater than zero to about 1.0 weight percent of calcium,from about 0.25 weight percent to about 1.0 weight percent of silver,from about 0.25 weight percent to about 1.0 weight percent of zirconium,and a balance of magnesium, based on total weight of the composition.

In one embodiment, the biodegradable, metal-alloy containing compositionincludes from about 0.5 weight percent to about 4.0 weight percent ofytrrium, from greater than zero to about 1.0 weight percent of calcium,from about 0.1 weight percent to about 1.0 weight percent of cerium,from about 0.25 weight percent to about 1.0 weight percent of zirconium,and a balance of magnesium, based on total weight of the composition.

In one embodiment, the biodegradable, metal-alloy containing compositionincludes from about 0.5 weight percent to about 4.0 weight percent ofytrrium, from greater than zero to about 1.0 weight percent of calcium,from about 0.25 weight percent to about 1.0 weight percent of silver,from about 0.1 weight percent to about 1.0 weight percent of cerium,from about 0.25 weight percent to about 1.0 weight percent of zirconium,and a balance of magnesium, based on total weight of the composition.

In one embodiment, the biodegradable, metal alloy-containing compositionincludes from about 1.0 to about 6.0 weight percent of zinc, from about0.25 to about 1 weight percent of silver, from greater than zero toabout 1.0 weight percent of zirconium, and a balance of magnesium, basedon total weight of the composition.

In one embodiment, the biodegradable, metal alloy-containing compositionincludes from about 1.0 to about 6.0 weight percent of zinc, from about0.1 to about 1 weight percent of cerium, from greater than zero to about1.0 weight percent of zirconium, and a balance of magnesium, based ontotal weight of the composition.

In one embodiment, the biodegradable, metal alloy-containing compositionincludes from about 1.0 to about 6.0 weight percent of zinc, from about0.25 to about 1 weight percent of silver, from about 0.1 to about 1weight percent of cerium, from greater than zero to about 1.0 weightpercent of zirconium, and a balance of magnesium, based on total weightof the composition.

In certain embodiments, the compositions of the invention are devoid ofzinc and aluminum. In another embodiment, the compositions of theinvention are devoid of aluminum. In still another embodiment, thecompositions of the invention may contain an amount of zinc and/or anamount of aluminum that is such as to maintain the toxicity levels ofthe compositions within acceptable limits. It is known that the presenceof zinc and/or aluminum in particular amounts can produce an undesirableor unacceptable level of toxicity in a physiological environment, suchas a body of a patient.

The biodegradable, metal alloy-containing compositions of the inventioncan be prepared using various methods and processes. In general, meltingand casting methods and processes are employed. It is known in the artof metallurgy that casting is a production technique in which a metal ora mixture of metals is heated until molten and then, poured into a mold,allowed to cool, and thereby solidify. In certain embodiments, themelted or molten metal or mixture of metals is poured into the mildsteel/copper mold at room temperature to 500° C.

Casting of the compositions of the invention can be affected by usingany casting procedure known in the art, such as, but not limited to,sand casting, gravity casting, permanent mold casting, direct chillcasting, centrifugal casting, low/high pressure die casting, squeezecasting, continuous casting, vacuum casting, plaster casting, lost foamcasting, investment casting, and lost wax casting. It is believed thatthe particular process used for casting can affect the properties andcharacteristics of the cast composition. Further, it is believed thatthe temperature at which the melting procedure is performed can alsoaffect the composition. Thus, the temperature may be carefully selectedso as to maintain the desired composition of the alloy.

In certain embodiments of the invention, yttrium, calcium, zirconium andmagnesium components (in specified amounts described herein) are meltedby heating at an elevated temperature, preferably under a protectiveatmosphere, and then poured into a mold, allowed to cool and solidify.In another embodiment of the invention, zinc, zirconium and magnesiumcomponents (in specified amounts described herein) are melted by heatingat an elevated temperature, preferably under a protective atmosphere,and then poured into a mold, allowed to cool and solidify.

In certain embodiments, prior to solidification, the molten mixture istested to determine the amount of the various components therein andtherefore, to provide an opportunity to adjust the amounts as desiredprior to solidification.

In other embodiments, the melting and/or casting steps are/is performedunder a protective atmosphere to preclude, minimize or reduceoxidation/decomposition of the components in the composition. Inparticular, it is desirable to preclude, minimize or reduce theoxidation/decomposition of magnesium in the composition. The protectiveatmosphere can include compounds selected from those known in the art,such as but not limited to, argon, sulfur hexafluoride, carbon dioxide,dry air and mixtures thereof.

In yet other embodiments, subsequent to the casting process, themagnesium-containing cast is subjected to homogenization. Withoutintending to be bound by any particular theory, it is believed that ahomogenization treatment can cause the spreading of, or more even oruniform distribution of, impurities, secondary phase(s), andinter-metallic phases, if present therein.

In further embodiments, the resulting cast can be subjected to variousforming and finishing processes known in the art. Non-limiting examplesof such processes include, but are not limited to, extrusion, forging,rolling, equal channel angular extrusion, stamping, deep-drawing,wire-drawing, polishing (by mechanical and/or chemical means), surfacetreating (to form a superficial layer on the surface) and combinationsthereof.

The resulting cast can be formed, finished, machined and manipulated toproduce articles and devices for use in medical applications, such asmedical devices for implantation into a body of a patient. Furthermore,these medical devices can be used in orthopedic, craniofacial andcardiovascular applications.

Detailed exemplary procedures for performing the melting and castingprocesses are depicted in the following examples.

The biodegradable, metal alloy-containing compositions of the inventioncan be used to produce various articles, such as medical devicessuitable for implantation into a body of a patient. In preferredembodiments, the medical implant devices include orthopedic,craniofacial and cardiovascular devices.

Additional objects, advantages and novel features of the invention maybecome apparent to one of ordinary skill in the art based on thefollowing examples, which are provided for illustrative purposes and arenot intended to be limiting.

EXAMPLES Example 1 1.1 Material Preparation

Ingots of elemental magnesium (99.97% pure from U.S. Magnesium, Inc.),calcium (99.5% pure from Alfa-Aesar) and magnesium-yttrium master alloy(4 wt. % yttrium from GKSS in Germany) were weighed according to thenominal composition. The ingots were melted together in a graphitecrucible (200 g batch) inside a quartz tube of a vacuum inductionfurnace to preclude oxidation of the pure elements. The graphitecrucible preloaded with batch and the quartz tube assembly were purgedwith UHP argon several times and vacuumed subsequently to achieve amoisture-free environment prior to induction melting. The inductionmelting then was conducted and repeated several times in order toachieve compositional homogeneity. The initial alloy produced by theinduction melting was cleaned thoroughly from any residue or oxide scaleand re-melted subsequently in a mild steel crucible using an electricalresistance furnace (from Wenesco, Inc.). The melting and pouringtemperature was about 700° C., and once the temperature was reached, anequivalent amount of zirconium was added using Zirmax® (Mg-33.3% Zr)master alloy (from Magnesium Elektron, LTD.). The liquid melt wasstirred for about 10 seconds after 1-minute and 5-minute intervals todissolve and disperse the zirconium particles uniformly into the melt.The melt was held for about 30 minutes at 700° C. and then poured onto acopper mold (1.5″×0.5″) and a steel mold (2.0″×1.5″) at roomtemperature. The as-cast samples were solution treated (“T4”) at 525° C.for about 2 hours inside a tubular furnace covered with magnesiumgettered powder under a protective atmosphere of argon and sulfurhexafluoride, and then quenched into water. Thin square plates (10×10×1mm³) of samples were sectioned (using a Buehler Precision Saw Simplimet1000) from the as-cast and the T4 samples, and were characterized byX-ray diffraction (XRD) using Philips XPERT PRO system employing theCuKα (λ=1.54056 Å) radiation operated at 45 kV and 40 mA to determinethe phase evolution and formation. The thin plate samples from theas-cast and T4 conditions were also used for electrochemical corrosion,cytotoxicity and cell adhesion tests. Each square plate sample wasmechanically grinded and polished to 2000 grit; ultrasonically cleanedin acetone, absolute ethanol and distilled water; and then dried in avacuum oven at a temperature of 50° C. For cytotoxicity tests, sampleswere sterilized by ultraviolet radiation for about 1 hour.

1.2 Cytotoxicity Test

A murine osteoblastic cell line (MC3T3-E1) was obtained from AmericanType Culture Collection (“ATCC”, Rockville, Md.) and used in the invitro experiment to determine the viability of cell attachment to WXK 10alloys. The cells were cultured in Modified Eagle's Medium alpha (αMEM),10% Fetal Bovine Serum (FBS), 100 U ml⁻¹ penicillin and 100 μg ml⁻¹streptomycin, and incubated at a temperature of 37° C. in a humidifiedatmosphere with 5% CO₂. The alloy samples were incubated in MEM forabout 10 minutes after which the cells were seeded on the as-cast and T4samples, as well as, as-rolled AZ31 control samples, at a cell densityof 4×10⁴ cells/well. After 24 hours of culturing at 37° C. in ahumidified atmosphere with 5% CO₂. the media was removed and thelive/dead cell viability assay was performed using a commerciallyavailable kit (obtained from Invitrogen Corporation. Karlsruhe,Germany). This kit was designed to determine the viability/cytotoxicityof cells by differentiating between live and dead cells withfluorescence microscopy of two different colors. The live/dead solutionwas composed of PBS, ethidium homodimer-1 (EthD-1) and calcein AM. Afterincubation in the live/dead solution for about 30 minutes at roomtemperature, images of the live and dead cells were captured usingfluorescence microscopy. The excitation wavelength of 495 nm was usedfor the fluorescence imaging microscopy. The live cells were observed asgreen (515 nm) fluorescent by enzymatic conversion of calcein AM tolabeled calcein. The dead cells were displayed as red (635 nm) byfluorescence enhancement upon entering and binding EthD-1 to nucleicacid due to low membrane integrity.

1.3 Direct Cell Adhesion Test

Following the live/dead cytotoxicity test, the samples were rinsed inPhosphate Buffer Solution (PBS, pH=7.4), fixed in 2.5% glutaraldehydesolution for about 15 minutes at room temperature, rinsed 3 times withthe PBS, followed by dehydration in a gradient ethanol/PBS mixture (30%,50%, 70%, 90%, 95%, 100%) for about 10 minutes each and then, dried. Thesurface of the cell attached samples were observed using Philips XL-30FEG scanning electron microscopy (SEM).

1.4 Results

A XRD pattern was generated of the Mg-1% Y-0.6% Ca-0.4% Zr alloy (WXK11)cast into two different molds (i.e. Cu mold and steel mold). It wasevident from the XRD pattern that only the α-Mg phase formed duringsolidification. During alloy design, the alloying additions (Y, Ca) werechosen within their maximum solid solubility limits according to theestablished phase diagrams to minimize the microgalvanic corrosion pathin the biological environment of αDMEM primarily between the matrix andthe secondary phase(s). The maximum solid solubility of calcium is 1.12wt % at 517° C. and for yttrium, 11.4 wt % at 567.4° C. The XRD patternclearly showed the formation of α-Mg without any traces ofintermetallics during solidification of the liquid melt from the pouringtemperature (700° C.). Similarly, XRD analysis of the samples which weresolution treated at an elevated temperature (525° C.) for about 2 hoursalso confirmed the absence of any secondary phase(s) and predominantly,the XRD line patterns were indexed with α-Mg.

The osteoblastic MC3T3-E1 cells were cultured in direct αMEM for 24hours and then, stained with calcein-AM and EthD-1. It was evident uponobservation that the number of cells cultured in the negative controlcell culture dish and the rolled AZ31 plates remained live after about24 hours with few dead cells stained in red. However, the cell densitydecreased in rapid manner in the AZ31 plates which suggested that ionicdissolution, e.g. of Mg²⁺ and Ca²⁺, likely started early with a slightincrease in pH value recorded from 7.5 to 8. The WXK10 samples whichwere cast into two different molds showed improved results when comparedwith AZ31 as the cell density was more evenly distributed. The solutiontreated (T4) sample showed an appreciable increase in the cell densitycovering most of the surface as compared to the as-cast samples. Therewas no significant difference in the shape of the viable cells (green)between the control and the studied samples groups. Only a few apoptoticcells (red fluorescence bound to nucleic acids) were seen in each group.

Example 2 2.1 Material Preparation

Ingots of elemental magnesium (99.97% pure from U.S. Magnesium, Inc.),zinc (99.99% pure from Alfa-Aesar) were melted together in a mild steelcrucible inside an electrical resistance furnace (Wenesco Inc.). Atypical melt size was 200 g. The melt was covered with a protective gasatmosphere (0.5% SF₆ with the balance Ar) to prevent magnesium burning.Once the desired pouring temperature (700° C.) was reached, anequivalent amount of zirconium was added using Zirmax® (Mg-33.3% Zr)master alloy (from Magnesium Elektron, LTD.). The liquid melt wasstirred for about 10 seconds after 1-minute and 5-minute intervals todissolve and disperse the zirconium particles uniformly into the melt.The melt was held for about 30 minutes at 700° C. and then poured onto acopper mold (1.5″×0.5″) at room temperature. The as-cast samples weresolution treated (“T4”) at 350° C. for about 1 hour inside a tubularfurnace under a protective atmosphere of gettered argon and sulfurhexafluoride, and then quenched into water. Thin square plates (10×10×1mm³) of samples were sectioned (using a Buehler Precision Saw Simplimet®1000) from the as-cast and the T4 samples, and were characterized byx-ray diffraction (XRD) using Philips XPERT PRO system employing theCuK® (λ=1.54056 Å) radiation operated at 45 kV and 40 mA to determinethe phase evolution and formation. The thin plate samples from theas-cast and T4 conditions were also used for electrochemical corrosion,cytotoxicity and cell adhesion tests. Each square plate sample wasmechanically grinded and polished to 2000 grit; ultrasonically cleanedin acetone, absolute ethanol and distilled water; and then dried in avacuum oven at a temperature of 50° C. For cytotoxicity tests, sampleswere sterilized by ultraviolet radiation for about 1 hour.

2.2 Cytotoxicity Test

A murine osteoblastic cell line (MC3T3-E1) was obtained from AmericanType Culture Collection (“ATCC”, Rockville, Md.) and used in the invitro experiment to determine the viability of cell attachment to ZK40alloys. The cells were cultured in Modified Eagle's Medium alpha (αMEM),10% Fetal Bovine Serum (FBS), 100 U ml⁻¹ penicillin and 100 μg ml⁻¹streptomycin, and incubated at a temperature of 37° C. in a humidifiedatmosphere with 5% CO₂. The alloy samples were incubated in αMEM forabout 10 minutes after which the cells were seeded on the as-cast and T4samples, as well as, as-rolled AZ31 control samples, at a cell densityof 4×10⁴ cells/well. After 24 hours of culturing at 37° C. in ahumidified atmosphere with 5% CO₂, the media was removed and thelive/dead cell viability assay was performed using a commerciallyavailable kit (obtained from Invitrogen Corporation, Karlsruhe,Germany). This kit was designed to determine the viability/cytotoxicityof cells by differentiating between live and dead cells withfluorescence microscopy of two different colors. The live/dead solutionwas composed of PBS, ethidium homodimer-1 and calcein AM. Afterincubation in the live/dead solution for about 30 minutes at roomtemperature, images of the live and dead cells were captured usingfluorescence microscopy. The excitation wavelength of 495 nm was usedfor the fluorescence imaging microscopy. The live cells were observed asgreen (515 nm) fluorescent by enzymatic conversion of calcein AM tolabeled calcein. The dead cells were displayed as red (635 nm) byfluorescence enhancement upon entering and binding EthD-1 to nucleicacid due to low membrane integrity.

2.3 Direct Cell Adhesion Test

Following the live/dead cytotoxicity test, the samples were rinsed inPhosphate Buffer Solution (PBS, pH=7.4), fixed in 2.5% glutaraldehydesolution for about 15 minutes at room temperature, rinsed 3 times withthe PBS, followed by dehydration in a gradient ethanol/PBS mixture (30%,50%, 70%, 90%, 95%, 100%) for about 10 minutes each and then, dried. Thesurface of the cell attached samples were observed using Philips XL-30FEG scanning electron microscopy (SEM).

2.4 Results

A XRD pattern was generated of the Mg-4% Zn-0.5% Zr alloy (ZK40) castinto a copper mold. It was evident from the XRD pattern that only theα-Mg phase formed during solidification. The amount of zinc added waswithin the maximum solubility limit of zinc, i.e., 6.2 wt % at atemperature of 341° C. according to the accepted phase diagram. The zincdissolved into the α-Mg lattice increased the solid-solutionstrengthening of the alloy.

The osteoblastic MC3T3-E1 cells were cultured in direct αMEM for 24hours and then stained with calcein-AM and EthD-1. It was evident thatthe number of cells cultured in the negative control cell culture dishand the rolled AZ31 plates remained live after about 24 hours with fewdead cells stained in red. However, the cell density decreased in rapidmanner in the AZ31 plates which suggested that ionic dissolution, e.g.,of Mg²⁺ and Ca²⁺, likely started early with a slight increase in pHvalue recorded from 7.5 to 8. The ZK40 sample which was cast into thecopper mold, as well as the heat-treated one (300° C., 1 hour) showedimproved results when compared with AZ31 as the cell density was moreevenly distributed. There was no significant difference in the shape ofthe viable cells (green) between the control and the studied samplesgroups. Only a few apoptotic cells (red fluorescence bound to nucleicacids) were seen in each group.

The morphology of the MC3T3-E1 cells was observed at differentmagnifications (100×, 200×, 1000×, 2000×) after 24 hour incubation inthe αMEM medium after fixing the cells in 2.5% glutaraldehyde solutionfor about 15 minutes. The cells were attached to the surface of thesample and it was evident that cells started growing. The cell spreadingwas uniform with filopodium and lammelipodium formations which suggestedthat the as-cast sample was stable in the bio-corrosive environment forcell growth and proliferation.

Example 3

In this example, yttrium (Y), calcium (Ca), zinc (Zn), silver (Ag) andzirconium (Zr) were alloyed in solid solution with magnesium (Mg) tocreate new Mg alloys. It is believed that Y contributed to grainboundary strengthening of the magnesium alloys as well as improvingcorrosion resistance with Y content above 3%, Ca improved corrosionresistance and mechanical properties of pure Mg up to 1 wt % Ca. Silver(Ag) provided anti-microbial properties, and Zr served as an effectivegrain refining agent, imparting grain boundary strengthening andcorrosion resistance. Density functional theory has shown alloying withCa and Y help to form a stable and chemically less reactive hydroxidelayer to impart greater corrosion resistance. The alloys in thisexample, Mg-1Y-0.6Ca-0.4Zr (wt. %), denoted henceforth as WXK11(codified according to ASTM B275-05), and Mg-4Y-0.6Ca-0.4Zr (wt. %),denoted henceforth as WXK41, were assessed based on theirbiocompatibility, corrosion behavior, and mechanical properties with theobjective of use in orthopedic medical implants. Biocompatibility wasdetermined in vitro using direct and indirect cell viability tests.Corrosion behavior was evaluated electrochemically and using hydrogenevolution. Mechanical properties were measured by both compressing andtensile loading. The novel alloys were compared in their as-cast and T4solution heat treated conditions, exhibiting improved biocompatibility,corrosion resistance, and mechanical properties as compared to pure Mg.

3.1 Material Preparation and Characterization

Novel magnesium, Mg-based polycrystalline, amorphous alloys weredeveloped using conventional gravity/permanent mold casting, high energymechanical milling, powder metallurgy and pulsed laser depositiontechnique. The alloying elements (Zn, Ca, Y, Ce, Ag, Zr, Al, MnSr) werecarefully selected based on the first principle theoretical calculationusing Vienna ab-initio Simulation Package (VASP) and composition wasselected keeping the constituent solute elements (Zn, Ca, Y, Ce, Ag, Zr,Al, Mn, Sr) within the maximum solid solubility (Cs) limit at theliquidus temperature (T_(L)) of the established phase diagrams to impartan equiaxed microstructure. The following compositions were explored indeveloping novel polycrystalline magnesium alloys: ZK series: Mg-1-6%Zn-0.25-1% Zr, ZQK series: Mg-1-6% Zn-0.1-1% Ag-0.25-1% Zr, ZQEK series:Mg-1-6% Zn-0.1-1% Ag-0.1%-1% Ce-0.25-1% Zr, WXK series: Mg-1-4% Y-0.3-1%Ca-0.25-1% Zr, WXQK series: Mg-1-4% Y-0.3-1% Ca-0.1-1% Ag-0.25-1% Zr,WXEK series: Mg-1-4% Y-0.3-1% Ca-0.1-1% Ce-0.25-1% Zr, WXQEK series:Mg-1-4% Y-0.3-1% Ca-0.1-1% Ag-0.1-1% Ce-0.25-1% Zr, AZXM series: Mg-1-9%Al-0.5-6% Zn-0.3-1% Ca-0.1-1% Mn, AZXMQ series: Mg-1-9% Al-0.5-6%Zn-0.3-1% Ca-0.1-1% Mn-0.1-1% Ag, AZXMW series: Mg-1-9% Al-0.5-6%Zn-0.3-1% Ca-0.1-1% Mn-1-4% Y, AZXMEseries: Mg-1-9% Al-0.5-6% Zn-0.3-1%Ca-0.1-1% Mn-0.1-1% Ce JX series: Mg-1-4% Sr-0.3-1% Ca alloys, JZseries: Mg-1-4% Sr-1-6% Zn alloys, JZX series: Mg-1-4% Sr-1-6% Zn-0.3-1%Ca alloys, JZQX: Mg-1-4% Sr-1-64% Zn 0.1-1% Ag-0.3-1% Ca alloys,JZXQEseries: Mg-1-4% Sr-1-6% Zn0.1-1% Ag-0.3-1% Ca-0.1-1% Ce, JZXQW:Mg-1-4% Sr-1-6% Zn0.1-1% Ag-0.3-1% Ca1-4% Y

Pure elemental ingots of Mg (US Magnesium Inc., Salt Lake City, Utah,99.97%), Ca shots (Alfa-Aesar, Ward Hill, Mass., 99.5%). Zn granules(Alfa-Aesar 99.99%), Al shots (Alfa-Aesar 99.99%). Mn shots (Alfa-Aesar99.9%), Ag (Alfa-Aesar 99.95%), Mg-5 wt % Ce master alloy, and Mg-4 wt %Y master alloy (Helmholtz-Zentrum Geesthacht Centre for Materials andCoastal Research, Germany) at varying compositions discussed above wereweighed according to the nominal composition and melted together in agraphite crucible inside an induction furnace (MTI Corporation,Richmond, Calif.), purged with ultrahigh-purity Ar and vacuumed to avoidoxidation of the pure elements. The initial alloy produced by inductionmelting was cleaned thoroughly from any residue or oxide scale andre-melted subsequently in a mild steel crucible using an electricalresistance furnace (Wenesco Inc., Chicago, Ill.) under the protection ofAr+1.5% SF₆ cover gas. The melting and pouring temperature was between700-850° C., and once the temperature was reached, an equivalent amountof zirconium was added using Zirmax® (Mg-33.3% Zr) master alloy(Magnesium Elektron Ltd., Manchester, UK). The melt was stirred and heldfor 30-60 minutes and then poured into a cylindrical steel moldpreheated to a temperature of 300-500° C. with an inner diameter of 44mm. The as-cast samples were solution treated (T4) at a temperature of300-550° C. for a period of 2-24 hours inside a tubular furnace coveredunder continuous Ar flow and quenched in water. A few selected alloyswere also artificially aged (T6 treatment) in an oil bath between atemperature of 150-300° C. for a period of 12-72 hours. The alloynominal compositions, determined by inductively coupled plasma opticalemission spectroscopy (ICP-OES, iCAP duo 6500 Thermo Fisher, Waltham.MA), are listed in Table 1.

TABLE 1 Chemical Composition Obtained from ICP-AES Analysis ofMg—Y—Ca—Zr, Mg—Zn—Zr, and Mg—Y—Ca—Ag—Zr Alloys (wt. %) Chemicalcompositions (wt. %) Alloy Y Ca Zr Cu Fe Mn Ni Si Mg (WXK11) 0.66 ± 0.030.52 ± .01  0.13 ± 0.004 0.016 0.003 0.008 0.008 0.006 BalanceMg—1Y—0.6Ca—0.4Zr (WXK41) 3.28 ± 0.001 0.42 ± 0.002 0.08 ± 0.001 0.0150.014 0.006 0.003 0.007 Balance Mg—4Y—0.6Ca—0.4Zr Chemical compositions(wt. %) Alloy Zn Zr Cu Fe Mn Ni Si Mg (ZK40) 4.28 ± 0.11 0.36 ± 0.0080.014 0.002 0.003 0.018 0.007 Balance Mg—4Zn—0.5Zr Chemical compositions(wt. %) Alloy Y Ca Ag Zr Cu Fe Mn Ni Mg (WXQK11) 0.51 ± 0.017 0.49 ±.022 0.21 ± 0.005 0.182 ± 0.018 0.014 0.004 0.008 0.010 BalanceMg—1Y—0.6Ca—0.25Ag—0.4Zr

Square plate samples (10×10×1 mm³) were sectioned using a diamond saw(Precision Saw Simplimet 1000, Buehler, Lake Bluff, Ill.) from theas-cast and the T4 samples for phase and microstructurecharacterization, electrochemical corrosion, and direct in vitro cellculture studies. Rod samples of 6 mm diameter and 6 mm length weremachined indirect in vitro cell studies. Rod samples of 10 mm diameterand 20 mm length were machined for compressive tests. As-cast Mg (USMagnesium, Inc.) was used as a comparison.

Phase characterization was conducted by X-ray diffraction (XRD) usingPhilips X'Pert PRO diffractometer employing CuK_(α) (λ=1.54056 Å)radiation with a Si-detector (X'celerator). The X-ray generator operatedat 45 kV and 40 mA at a 20 range of 10-90°. Samples were mechanicallyground and polished up to 1200) grit, ultrasonically cleaned inisopropyl alcohol, and air dried. For cytotoxicity tests, samples weresterilized by ultraviolet radiation for 1 hour.

3.2 Microstructure Characterization

Square plate samples of the ZK, WXK, WXQK alloy series were mounted inepoxy (EpoxiCure, Buehler), mechanically polished (Tegramin-20, Struers,Ballerup, Denmark), and chemically etched in a solution of 5 mL aceticacid, 6 g picric acid, 10 mL water, and 100 mL ethanol. Themicrostructure was observed under an optical microscope (Axiovert 40MAT, Carl Zeiss, Jena, Germany).

3.3 Mechanical Properties

Samples were machined along the long axis of the various alloy ingots inaccordance with ASTM-E8-04 for tensile testing and ASTM-E9-09 forcompressive testing. Sample dimensions for tensile and compressivetesting were as follows: standard dogbone specimens (tensilemeasurements: gauze length: 12.3 mm, gauze cross-section: 3 mm×3 mm);(compressive measurements: 10 mm dia×20 mm length). Tensile andcompressive stress-strain curves were obtained for as-cast and T4solution treated alloys, and compared to as-cast pure Mg. The tensileand compressive tests were conducted at room temperature at a cross-headspeed of 2 mm/min using an Instron universal testing system with laserextensometer by OrthoKinetic® Testing Technologies. Yield strength (YS),Ultimate Tensile Strength (UTS), Young's modulus (E) during compressionand tension, percent elongation (%), compressive yield strength,compressive peak strength, percent compression of various alloys wasdetermined from the stress-strain curves. The tensile and compressiveyield strengths were determined from the linear portion of thestress-strain curve during the tensile and compressive tests.

3.4 Electrochemical Corrosion Test

To test corrosion of the ZK, WXK, WXQK alloys, the potentiodynamicpolarization technique was used. Samples were connected to a copper wireusing silver epoxy and mounted in epoxy resin. The mounted samples weremechanically polished, sonicated in isopropyl alcohol, and dried in air.The potentiodynamic corrosion study was carried out with anelectrochemical workstation (CH-604A, CH Instruments, Inc., Austin,Tex.) at a scanning rate of 1 mV/s and potential window of 500 mV aboveand below the open circuit potential. A three electrode cell wasemployed with platinum as the counter electrode. Ag/AgCl as thereference electrode, and the sample mounted in epoxy resin as theworking electrode. The test was performed in Dulbecco's Modified EagleMedium (DMEM, with 4.5 g/L glucose, L-glutamine, and sodium pyruvate,Cellgro, Manassas, Va.) supplemented with 10% fetal bovine serum (FBS)at pH 7.2±0.2 and held at 37.4° C. Before each measurement, the samplewas immersed in DMEM to provide stability. The cathodic and anodicportions of the generated Tafel plots were fit linearly to allowcalculation of corrosion potential, E_(corr) and corrosion currentdensity, i_(corr). Samples were cleaned by immersion in 200 g/L ofchromic acid and 10 g/L of AgNO₃ for 10 minutes to remove corrosionproducts and corrosion morphology was characterized using SEM and EDX.

3.5 Immersion Corrosion Test (Weight Loss)

Immersion tests were carried out in conformation with ASTM G31-72 (theratio of surface area to solution volume was 1 cm²:20 ml). Samples wereremoved after 1 and 3 weeks of immersion, rinsed with distilled waterand dried at room temperature. The samples were cleaned by immersion in200 g/L of chromic acid and 10 g/L of AgNO₃ for 10 minutes to removecorrosion products and the degradation rates (in units of mm/year) wereobtained according to ASTM-G31-72. The corrosion rate is given by Eq.(1):

Corrosion rate=(K×W)/(A×T×D)  Eq. (1)

wherein the coefficient K=8.76×10⁴, W is the weight loss (g), A is thesample area exposed to solution (cm²), T is the exposure time (h) and Dis the density of the material

(g cm⁻³). The pH value of the solution was also recorded during theimmersion tests.

3.6 Indirect Cytotoxicity Tests

ZK, WXK, WXQK alloy samples and as-cast pure magnesium were polished upto 1200 grit, ultrasonically cleaned in isopropyl alcohol, air dried,and sterilized by ultraviolet radiation for 1 hour. The specimens wereincubated in modified Eagle's medium alpha (αMEM) supplemented with 10%fetal bovine serum (FBS), 100 U/ml penicillin, and 100 μg/mlstreptomycin at a temperature of 37° C. in a humidified atmosphere with5% CO₂ for a period of 72 hours. The sample weight to extraction mediumratio was 0.2 g/mL in accordance with the EN ISO standard 10933:12. Thisextraction ratio was designated as 100% extract, with less concentratedextracts prepared by diluting the 100% extract into 50%, 25%, and 10%extract solutions. Extracts were sterile filtered using 0.2 μm syringefilter before being added to cells.

The murine osteoblastic cell line (MC3T3-E1, American Type CultureCollection, Rockville, Md.) was used in in vitro cell cytotoxicityexperiments, cultured in modified Eagle's medium alpha (αMEM), 10% fetalbovine serum, 100 U/ml penicillin and 100 μg/ml streptomycin at atemperature of 37° C. in a humidified atmosphere with 5% CO₂. The cellswere seeded in 96-well cell culture plates at 6×10³ cells/200 μl mediumin each well and incubated for 24 hours to attach before adding theextraction medium. The controls used culture medium without extract asthe negative control and 10% DMSO culture medium as the positivecontrol. The medium was then replaced with 200 μl of extraction mediumat 100%, 50%, 25%, and 10% extract concentrations and incubated undercell culture conditions for 3 days. The cytotoxicity of the corrosionextracts were tested using the MTT assay. Media and extracts werereplaced with fresh cell culture medium to prevent interference of themagnesium in the extract from interacting with the tetrazolium salt. TheMTT assay was performed according to the Vybrant MTT Cell ProliferationKit (Invitrogen Corporation, Karlsruhe, Germany) by first adding 10 μlof 12 mM 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) dissolved in phosphate buffer solution (PBS, pH=7.4) to each well.The samples were incubated at a temperature of 37° C. with MTT for 4hours, after which 100 μl formazan solubilization solution (SDS-HClsolution) was added to each well and incubated for 12 hours. Theabsorbance of the samples was measured using the Synergy 2 Multi-ModeMicroplate Reader (BioTek Instruments, Winooski, Vt.) at a wavelength of570 nm. The absorbance of the samples was divided by the absorbance ofthe mean positive control subtracted from the mean negative control todetermine percent viability of cells compared to the controls.

3.7 Direct Cell Viability and Adhesion Test

MC3T3-E1 cells were cultured directly on ZK40, WXK11, WXK41, WXQK11alloys and as-cast pure magnesium. Cell culture conditions and mediawere the same as in the indirect cytotoxicity test. Samples were cut todimensions of 10 mm×10 mm×1 mm and polished up to 1200 grit,ultrasonically cleaned in acetone, air dried, and sterilized byultraviolet radiation for 1 hour. The alloy samples were incubated inαMEM with 10% fetal bovine serum (FBS), 100 U/ml penicillin, and 100μg/ml streptomycin for 10 minutes after which cells were seeded on thesamples at a cell density of 4×10⁴ cells/mL. Viability of the seededcells was evaluated at 1 and 3 days using the LIVE/DEADViability/Cytotoxicity Kit (Invitrogen Corporation, Karlsruhe, Germany)following manufacturer protocol. This kit determines theviability/cytotoxicity of cells by differentiating between live and deadcells with fluorescence microscopy of two different colors. Briefly, thealloy samples with attached MC3T3-E1 cells were washed with PBS andstained for 30 minutes at room temperature with 2 μmol/L ethidiumhomodimer-1 and 4 μmol/L calcein AM in PBS. After incubation in theLive/Dead solution for 30 minutes in room temperature, live and deadcells images were captured using fluorescence microscopy.

3.8 Results

XRD patterns of the ZK40, WXK11, WXK41 and WXQK11 in as-cast andsolution treated (T4) condition were generated. The XRD patterns clearlyshowed all the alloys were composed of α-Mg with an hcp crystalstructure, without the presence of unalloyed Y, Ca, Zn, Zr and otherintermetallic compounds. The XRD patterns clearly showed that only α-Mgsolid solution single phase was formed during solidification in thefinal microstructures. During alloy design, the alloying elements e.g.,(Y, Ca, Zn, Zr) were carefully selected within the solid solubilitylimits at the liquidus temperature (T) of the phase diagrams with theconsideration that the final microstructure should be free of any 2^(nd)phase/intermetallic phases along the grain boundary regions which arehighly cathodic in nature and accelerate microgalvanic corrosion in asuitable physiological environment primarily between the matrix and thesecondary phase(s). Chemical composition of each of the alloys wasdetermined by ICP-AES analysis. Table 1 showed that the actualcomposition of each alloy was within its nominal composition. However,slight decrease in the yttrium, calcium, and zinc was likely due to are-melting process. It was believed that the loss in total zirconiumcontent was primarily due to settling of large zirconium particles andclusters in the liquid melt. The impurity content of each of the alloycompositions was very low ensuring improved biocompatibility anddegradation properties.

Optical micrographs of ZK40, WXQK11, WXK11 and WXK41 alloys in theas-cast and solution treated conditions were generated. The grain sizewas calculated using a linear-intercept method [ASTM E112]. The averagegrain size of ZXK40, WXQK11, WXK11, WXK41 alloys in as-cast conditionwere 60 μm, 130 μm, 110 μm, 80 μm, respectively, with the presence ofuniform equiaxed α-Mg grains throughout the microstructure. However, theslight presence of secondary phase(s) was also evident along the grainboundary due to second phase(s) precipitates during solidification whichis a common phenomenon during casting. The WXK as-cast ingot sampleswere subjected to solution heat treatment at elevated temperature (525°C.-6 hours) followed by quenching in water to impart a more uniform,homogeneous microstructure. The microstructure after T4 treatment ofWXK11, WXK41 alloys showed that there was a slight increase in grainsize likely due to coalescence of smaller grains together along thetriple point grain boundary regions and formation of supersaturated α-Mggrains after the precipitates dissolved into the matrix.

Table 2 summarizes the mechanical properties of the as-cast and T4treated alloys herein as compared with commercial AZ31 and pure Mg.Table 2 shows the Young's modulus of the new alloys (E ˜64 GPa for ZK40,51 GPa for WXK11, 38 GPa for WXK41, 51 GPa for WXQK11) were comparablewith commercial AZ31 sheet (55 GPa) suggesting that the stiffness of thealloys are sufficient for orthopedic fixation and craniofacial andcardiovascular device applications. However, the new alloys demonstratedsurprisingly low value in the yield strength, and ultimate tensilestrength as compared to AZ31. It is believed that a potential reason forlow value in tensile strength was the presence of castingdefects/inclusion in the microstructure. In order to improve thestrength and ductility, the alloys were solution treated at elevatedtemperature and immediately quenched into water to improve the ductilityand better shape forming ability in expense of mechanical strength.Although there was a slight increase in elongation, a drastic drop inyield strength and tensile strength was evident.

TABLE 2 Mechanical Properties of As-Cast and T4 Treated Alloys UltimateCompres- Young's Yield Tensile sive Percent Modulus Strength StrengthStrength Elongation Alloy (GPa) (MPa) (MPa) (MPa) (%) Commercial 55 202268 409 12 AZ31 Pure Mg 5 19 66 180 7 ZK40 as- 64 96 176 363 4 cast ZK40T4 68 92 83 355 1.5 WXK11-as 51 72 123 296 3 cast WXK11-T4 49 45 106 2264 WXK41-as 38 89 162 306 6 cast WXK41-T4 34 44 83 227 3.5 WXQK11-as 5163 130 300 4 cast WXQK11-T4 33 45 114 284 6.6

The potentiodynamic corrosion behavior of the ZK40, WXK11, WXK41, andWXQK11 alloys in as-cast and solution treated condition along withas-cast pure Mg was studied extensively under physiological condition.The potentiodynamic polarization curves (Tafel plot) of the varioussamples and pure Mg, recorded at a scan rate of 1 mV/s in the presenceof DMEM, were plotted. The cathodic branch of the tafel plot showed thehydrogen evolution through a reduction process whereas the anodic branchrepresented the magnesium dissolution by oxidation. The cathodicplateaus of pure Mg suggested that the hydrogen evolution started at 1.7V. However, the calculation of corrosion current density, i_(corr)tabulated in Table 3, clearly showed that the corrosion current densityof the ZK40, WXK11, WXK41, and WXQK11 alloys were comparable to pure Mg(30.68 A cm⁻²) and commercial rolled AZ31 sheets (19.20 μA cm⁻²). Thecorrosion potential, E_(corr) of the ZK40, WXK11, WXK41, WXQK11 were 500mV higher compared to pure Mg which indicated that the samples were morestable in DMEM, due to formation of a protective film of corrosionproduct and subsequent passivation of the samples. One noticeabledifference was the decrease in the i_(corr) value of solution treatedsamples compared to as-cast samples (see Table 3) likely due toformation of supersaturated phase and reduction in the volume fractionof secondary phase(s) observed along the grain boundary which can act ascathodic sites for corrosion and also the presence of Zn, Y, Zr whichare able to elevate the corrosion potential in the anodic sites resultedimprove corrosion rate. The present corrosion study clearly showed thatthe current alloys are stable in aggressive physiological condition.

TABLE 3 Electrochemical Corrosion Measurements (Using Tafel Plots) Dataof Various Alloy Corrosion potential, Corrosion current CorrosionE_(corr) (V) vs. Ag/AgCl density, i_(corr) rate Material (R.E.) (μAcm⁻²) (mm/year) Pure Mg −1.62 30.68 0.70 Commercial −1.48 19.20 0.43AZ31 ZK40 as-cast −1.49 39.69 0.90 ZK40 T4 −1.55 39.32 0.87 WXK11as-cast −1.51 36.42 0.84 WXK11 T4 −1.41 5.70 0.13 WXK41 as-cast −1.5616.70 0.5 WXK41 T4 −1.54 5.22 0.12 WXQK11 as-cast −1.61 58.88 1.35

The SEM micrographs of corroded surface of the samples where thecorrosion product was cleaned with CrO₃/AgNO₃ solution were generated.It was evident from the SEM micrographs that corrosion was localized andpossibly occurred in the weak grain boundary region which is prone toattack under physiological condition. Formations of small localizedcavities throughout the microstructures clearly gave indication thatalloy purity and presence of secondary phase/defects are related tocontrolling and minimizing the degradation rate.

The immersion corrosion plot for ZK40 as-cast and solution treatedsamples, and for a period of 1 week and 3 weeks, respectively weregenerated. The corrosion rate was in good agreement with potentiodynamicpolarization data (Table 3). However, the exact reason for an increasein corrosion rate over a period of 3 weeks was not clear.

The indirect cytotoxicity results of ZK40 samples were obtained usingMC3T3-E1 cells and the MTT assay for 3 days extract. For both cultureperiods, cell viability was most reduced with 100% extractconcentration, and increased as the extract percentage decreased, withno cytotoxicity (>75% viability) observed at 50% or 25% extractconcentration. This was consistent with previous findings that showedhigh extract concentrations were highly cytotoxic and led to osmoticshock, suggesting that a 10-fold extract dilution be used for as-castmagnesium materials.

Cell viability was also studied for WXK11 and WXK41 samples for as-castand T4 condition with the 1 and 3 days culture time with extracts. After1 day of culture with extract, both WX11 and WX41 as-cast and T4 treatedalloys showed higher cell viability compared to pure Mg at 25% and 10%extract concentration; however, no difference between them could beobserved after 3 days of culture.

Osteoblastic MC3T3-E1 cells cultured in direct αMEM for 3 days and thenstained with calcein-AM and EthD-1 were obtained. Live cells convertedcalcein AM to green fluorescent calcein through intracellular esteraseactivity, while EthD-1 entered cells with compromised membranes where itbinded with nucleic acids and produced a bright red fluorescence. ZK40as-cast sample as well as a solution treated one (350° C.-1 h) showedencouraging results when compared with AZ31 as the cell density was moreand evenly distributed. There was no significant difference in the shapeof the viable cells (green) between the control and the studied samplegroups. Only a few apoptotic cells (red fluorescence in the nuclei) wereseen in each group. Morphology of the MC3T3-E1 cells after 3 daysincubation in the αMEM medium after fixing the cells in 2.5%glutaraldehyde solution for 15 minutes was generated. The cells wereattached on the surface of the sample and it was also evident cellproliferation was already started. The cell spreading was uniform withfilopodium and lammelipodium formations which suggested that the as-castsample was stable in the physiological environment for cell growth andproliferation.

Pre-osteoblast MC3T3-E1 cells were cultured directly on the WX11 andWX41 alloys for 1 and 3 days, and then stained with calcein-AM andethidium homodimer-1 (EthD-1). After 1 day of culture, both WX11 andWX41 T4 heat treated alloys demonstrated comparable live cell densitycompared to tissue culture plastic. Pure Mg and the as-cast WX11 andWX41 alloys showed reduced live cell density compared to tissue cultureplastic. The WX41 alloys appeared to show higher density of live cellscompared to WX11, possibly due to the higher Y content resulting in amore stable corrosion layer on the surface of the alloy. After 3 days ofculture, WX11 as-cast and T4 treated as well as the as-cast WX41 alloysdemonstrated much lower live cell attachment, consistent with the resultof the indirect cytotoxicity test. WX41 T4 treated alloy showedexcellent biocompatibility with high live cell attachment throughout thesurface of the alloy, far superior to pure Mg and the other WX alloys.The higher cell density on WX41 T4 treated alloy after 3 daysdemonstrated proliferation of the attached MC3T3-E1 cells.

1. A biodegradable, medical implant device comprising a metalalloy-containing composition, comprising: from about 0.5 weight percentto about 4.0 weight percent of yttrium; from greater than zero to about1.0 weight percent of calcium: from about 0.25 weight percent to about1.0 weight percent of zirconium; and a balance of magnesium, based ontotal weight of the composition.
 2. The composition of claim 1, whereinthe composition further comprises silver in an amount of from about 0.25weight percent to about 1.0 weight percent based on the total weight ofthe composition.
 3. The composition of claim 1, wherein the compositionfurther comprises cerium in an amount of from about 0.1 weight percentto about 1.0 weight percent based on the total weight of thecomposition.
 4. A biodegradable, medical implant device comprising ametal alloy-containing composition, comprising: from about 1.0 weightpercent to about 6.0 weight percent of zinc; from greater than zero toabout 1.0 weight percent of zirconium; and a balance of magnesium, basedon total weight of the composition.
 5. The composition of claim 4,wherein the composition further comprises silver in an amount of fromabout 0.25 weight percent to about 1.0 weight percent based on the totalweight of the composition.
 6. The composition of claim 4, wherein thecomposition further comprises cerium in an amount of from about 0.1weight percent to about 1.0 weight percent based on the total weight ofthe composition.
 7. A method of preparing a biodegradable, medicalimplant device comprising a metal alloy-containing composition,comprising: melting together at least one component selected from thegroup consisting of yttrium, calcium, zirconium, and zinc, withmagnesium to obtain a melted magnesium alloy mixture; and casting saidmelted magnesium alloy mixture to obtain said biodegradable, metalalloy-containing composition.
 8. The method of claim 7 wherein the atleast one component is from about 1.0 weight percent to about 6.0 weightpercent of zinc; and from greater than zero to about 1.0 weight percentof zirconium, and wherein a balance is magnesium.
 9. The method of claim7, wherein said composition further comprises from about 0.25 weightpercent to about 1.0 weight percent silver and from about 0.1 weightpercent to about 1.0 weight percent of cerium.
 10. (canceled) 11.(canceled)
 12. (canceled)
 13. The biodegradable, metal alloy-containingmedical implant device of claim 1, wherein the medical device is anorthopedic device.
 14. The biodegradable, metal alloy-containing medicalimplant device of claim 1, wherein the medical device is a craniofacialdevice.
 15. The biodegradable, metal alloy-containing medical implantdevice of claim 1, wherein the medical device is a cardiovasculardevice.
 16. The biodegradable, metal alloy-containing medical implantdevice of claim 1, wherein the medical device further comprises at leastone active substance selected from the group consisting of molecule,compound, complex, adduct, composite and combinations thereof.
 17. Thebiodegradable, metal alloy-containing medical implant device of claim16, wherein the at least one active substance is selected from the groupconsisting of bone growth promoting agents, drugs, proteins,antibiotics, antibodies, ligands, DNA, RNA, peptides, enzymes, vitamins,cells and combinations thereof.
 18. The method of claim 7 wherein the atleast one component is from about 0.5 weight percent to about 4.0 weightpercent of yttrium, from greater than zero to about 1.0 weight percentof calcium, and from about 0.25 weight percent to about 1.0 weightpercent of zirconium, and wherein a balance is magnesium.